Pulmonary fibrosis is a frequent and devastating complication of acute or indolent lung injury for which current therapy is effective in only 1/3 of cases. The fibroproliferative response after lung injury is characterized by matrix fibrin deposition and collagen accumulation. Several recent in vivo studies indicate that the serine protease inhibitor, plasminogen activator inhibitor (PAI-1) contributes to the fibrotic response. For example, in bleomycin-injured mice, PAI-1 is up- regulated in fibroproliferative lesional fibroblasts, and genetic deletion of PAI-1 confers protection from lung injury-induced fibrin and collagen deposition. These in vivo data provide strong evidence of a fibrogenic role for PAI-1, however, the regulatory pathways involved in fibroblast PAI-1 expression have yet to be elucidated. Furthermore, the proposed mechanism of PAI-1's in vivo effect through inhibition of alveolar fibrin clearance remains unproven. Our goal is to understand the mechanism of regulation of PAI-1 expression, and its role in the molecular events that result in pulmonary fibrosis. We have recently shown that fibroblasts up-regulate their expression of PAI-1 in response to fibrin D dimer, a plasmin-generated proteolytic fragment of fibrin that is abundant in fibroproliferative lesions. The work proposed herein will advance the field by defining the mechanism(s) by which D dimer increases PAI-1 expression in fibroblasts, and by determining the effect of in vivo manipulation of alveolar fibrinolysis on PAI-1 expression and on the fibrotic process. In the first two specific aims, we will determine the relative importance of changes in PAI-1 transcription rates and mRNA half-fife, and identify the critical cis and trans acting factors in basal and D dimer-stimulated PAI-1 transcription in fibroblasts. In specific aim 3, we will study effect of experimentally-induced alveolar fibrinolysis, with its attendant generation of D dimer, on lung PAI-1 expression and on the fibrotic response to bleomycin lung injury. We hope the knowledge of the molecular events in pulmonary fibrosis will lead to novel identifiable targets for therapeutic modalities aimed at reducing the fibroproliferative response to lung injury.